1. Field of the Invention
The present invention relates generally to the fields of genetic and chemical screening and diseases associated with protein misfolding. More particularly, it concerns the development of a yeast-based system that can be used to screen for substances that provide therapeutic value for various diseases associated with protein misfolding. Methods for performing genetic and chemical screens using the yeast systems of the invention are also provided. One major class of diseases benefited by the methods of the invention are the neurodegenerative diseases including Parkinson's disease, Alzheimer's disease, Huntington's disease and the like.
2. Description of Related Art
The correct folding of a protein is a key event to attain proper biological function. Correct folding leads to the characteristic conformation of a protein which determines protein activity, aggregation, degradation, and function. Several proteins are implicated in neurodegenerative diseases, such as Parkinson's disease (PD), transmissible spongiform encephalopathies (TSEs), Alzheimer's disease (AD), familial amyloid polyneuropathy (FAP), prion diseases, and Huntington's disease (HD), among several others. These proteins form abnormal aggregates due to alternative folding mechanisms. These misfolded protein aggregates form insoluble fibrils which are then deposited in tissues. Fibrillogenesis is the cause of various pathologies involving neuronal degeneration. Deposition of insoluble fibrils in tissues leads to formation of plaques and tangles and eventual cellular degeneration, as the pathology progresses. Despite a lack of amino acid sequence homology of the fibril forming proteins, the fibrils have several common morphological features. For example, some common morphological features of amyloid fibers, (formed by amyloid proteins), include a cross β-structure, similar sizes, display of green birrefringence upon staining with congo red when observed under polarized light, Thioflavin T binding, etc.
One example of a disease based on fibrillogenesis, is the pathology of amyloidosis which is defined by the deposition of amyloid fibrils into tissues and is typified by Alzheimer's Disease (AD). Systemic amyloidosis are characterized by amyloid deposition throughout the viscera. Animal amyloid is a complex material composed partly of protein fibrils. The protein that comprises these fibrils varies from disease to disease. β-Amyloid is one of these proteins which is involved in the pathological progression of AD.
In the case of Parkinson's disease (PD), dopaminergic neurons in the brain undergo selective neurodegeneration. A highly conserved pre-synaptic protein, α-synuclein, with unknown function has been implicated in PD. Two different point mutations in α-synuclein, A53T and A30P, are involved in autosomal dominant familial PD. It is likely that conformational changes in α-synuclein lead to the typical proteinaceous accumulation and fibrillogenesis characteristic of such diseases. Purified full-length α-synuclein can form fibrils similar to those found in Lewy Bodies (cytosolic inclusions) in PD. The mechanism of fibrillogenesis has not been described, although recent data indicate that α-synuclein aggregation follows a nucleation-elongation mechanism, as suggested for the other disease-related proteins.
It is well recognized in the art, that once fibrilloid deposits have formed, there is no known therapy or treatment which significantly dissolves such deposits in situ (U.S. Pat. No. 5,643,562). Consequently, strategies based on prevention of protein aggregation and fibril formation is a major goal in the therapy or prevention of diseases associated with protein misfolding such as neurodegenerative diseases and type 2 diabetes. Thus, there is a need in the art of a system where one can identify therapeutic agents for diseases associated with protein misfolding which may have their therapeutic effect due to being either regulators of protein folding, and/or inhibitors of protein aggregation, and/or preventors and/or inhibitors of the process of fibrillogenesis, or those that can have an entirely different and possibly unknown mechanism of action. Furthermore, there is need that such a system provide a rapid and cost-effective screening method that will allow the identification of agents useful in the treatment, prevention and cure of diseases associated with protein misfolding.